The present invention relates to a novel device for culturing cells using a first membrane forming a first bag, having a spacer material disposed within the first bag so as to maintain each side of the first bag at a predetermined distance from the opposing side; a second membrane forming a second bag containing therein the first bag, having a spacer material disposed within the second bag so as to maintain a predetermined distance between the interior surface of the second bag and the exterior surface of the first bag; a housing which encases the first and second bags; and three inlets and outlets disposed therein for passing a gas, cell culture and cell nutrient into selected chambers formed thereby.
There has been increased interest in recent years in methods for the culturing of in-vitro cells, especially mammalian cells. It is generally known that cells require both oxygen and nutrients to propagate and flourish. Recognizing such requirements a number of devices have been designed attempting to cultivate in-vitro cells in environments of oxygen and nutrients simultaneously.
U.S. Pat. No. 3,997,396 describes one method for the propagation of living cells in-vitro by attaching and growing cells on one side or surface of a hollow fiber membrane, and by passing oxygen through the membrane from the other side in contact with the cells and simultaneously incubating the cells in a nutrient medium. Another device is described in U.S. Pat. No. 3,948,732 which describes a replaceable, sterilizable, cell growth assembly comprising a chamber structure of gas permeable, liquid impermeable material having an inner surface to which cells are attachable. The chamber is of a tubing configuration and is disposed in a plurality of layers in stacked relation. The spacer structures between the layers of tubing define a plurality of flow passages between the layers and enable a gaseous environment to bath the majority of the external surface area of the tubing layers. The chamber also includes an inlet conduit for introducing a culture media for flow-thru the plural layers of the chamber structures and an outlet conduit for receiving culture media from the chamber structures, and coupling structures for detachably connecting the inlet conduit to a source of culture media and the outlet conduit to an appropriate culture media receptacle.
U.S. Pat. No. 4,225,671 describes a process for the in-vitro biosynthesis of hormones, especially insulin, by producing the cells in one or more cell culture spaces separated by semipermeable flat membranes such that there is at least one cell culture space surrounded by cell media (nutrient) and gas. U.S. Pat. No. 4,225,671 demonstrates multiple layering of gas, cell culture, and cell media spaces by connecting the gas and cell media in series with other gas and cell media spaces and also by connecting the cell culture spaces in parallel with other cell culture spaces. This is accomplished by means of flat plate type membranes being stacked one on top of the other for forming spaces therein.
The problem with the aforementioned devices is the amount of dead space between the membrane plates which result in some cells receiving less oxygen and nutrients than others. Moreover, they require numerous external fittings for the inlets and outlets which can result in leakage, loss of cells and increased capital cost. These systems are also bulky in size, i.e. they take up to much space in the laboratory, which requires increasing the means for pumping nutrient, gas and cells there-through thus generating undesirable increases in heat from the larger pumping means. Increases in heat is unacceptable in culturing of mammalian cells or the like due to the pristine conditions required for propagation of the cells.
The present inventors developed an apparatus which overcomes the aforementioned channel spacing and dead space problems, as well as providing a cell culturing apparatus of reduced size and complexity.